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Whole –genome sequencing and the etiology of osteoporosis

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Osteoporosis is a disease with a strong genetic etiology (1). However, exactly which or how many genes are involved in the pathogenesis of this condition is unknown. Numerous studies have tried to link specific regions of the genome to traits such as bone mineral density to understand the genetic variations that predispose individuals to osteoporosis. In the past these have either used a genome wide association study (GWAS) approach or a candidate gene approach. GWAS uses single nucleotide polymorphisms (SNPs) or variations in DNA, which occur naturally throughout the genome, to identify regions of chromosomes that predispose individuals to have a specific trait such as high or low bone mineral density. The candidate gene approach examines specific variations in the DNA sequence of known genes (including the regulatory, coding and non-coding regions) to identify variations that occur more frequently with a specific trait like bone mineral density. Both approaches have their strengths and weaknesses. GWAS can identify regions in the entire genome that “link” to specific traits but not specific sequences in DNA that are causative for the development of disease. Candidate gene studies can identify specific sequences in a gene that are involved in the development of a trait but are limited to analysis of the sequences of known genes.


In contrast, whole genome sequencing uses the genetic information of the entire genome (approximately 3 billon base pairs from the DNA of the chromosomes and the mitochondria) to link specific sequence variations to human traits. The technology has evolved since 2000 when the Human Genome Project first published its rough draft at a cost of over 3 billion dollars. Currently, the cost of sequencing a human genome is approaching $1000 for a high fidelity run and it is likely that this price will drop further as the technology evolves. Reports that use whole genome sequencing to link human genetic sequence variations to bone density have now been published. The first was in 2013 and identified a mutation in the leucine-rich repeat-containing G-protein coupled receptor 4 (LGR4) gene that fully disrupts its function (2). This variation was identified in an Icelandic population and appears specific for this group since it was not detected in other European populations. More recently, a large international study (3) used the UK10K project to examine whole genome sequencing of 2,882 subjects as well as whole exome (the protein coding regions of genes) sequencing in 3,549 subjects and identified a non-coding region genetic variant in the vicinity of the engrailed homeobox-1 (EN1) gene, which associated with bone mass. This variant was relatively rare, with a minor allele frequency of 1.7% and is not represented in the current HapMap imputation panels, showing that there is still much to be learned about genetic variation in humans. Additional, albeit slightly less significant, variants around the EN1 gene were also found and these too are very rare (occurring in 1.6% to 5.8% of the population). Three of the four genetic variants associated with lumbar bone density while the fourth associated with femoral bone density. All these results demonstrate that bone mass is a complicated genetic phenomenon.


Unlike the previous Icelandic study, EN1 variants were present and segregated similarly with bone mass in the general population of subjects of European descent. Demonstrating the power of this technology, this observed effect was highly significant and fourfold larger than that previously seen at any chromosomal site in a large GWAS analysis. In addition, one of the DNA sequence variants was associated with increased bone mass and a decreased fracture risk in a large cohort study.


The authors also characterized a mouse model, which had a targeted deletion of En1. This animal had decreased bone mass with increased osteoclastic and osteoblastic activity. Additional studies found that En1 was expressed in osteoblast lineage cells but not in osteoclasts. Based on the results of all these studies, the authors concluded that En1 is an important mediator of skeletal biology.


These reports demonstrate the potential power of whole genome sequencing to identify relatively rare variants in the genome that associates with skeletal disease. However, they also demonstrate the complexity of such studies, which require a large number of subjects, sophisticated statistical genomics and advanced genome sequencing techniques. Because of these limitations, progress in identifying the multi-gene variations that may underlie more common causes of osteoporosis are not yet within reach. However, one only has to examine the rapidity with which the technology of whole genome sequencing has advanced in the last 15 years to appreciate that these challenges are apt to be overcome. We are most likely on the verge of important new insights into the genetic etiology of this disease.



Joe Lorenzo


Farmington, CT, USA

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